Lignin is the second most abundant organic material in the biosphere, and is a major component of cell walls of woody plants (such as poplar and pine species) and fodder crops (such as maize, wheat and barley). The quantity of lignin in plant material affects characteristics that are agronomically important. For example, in fodder crops the amount of lignin present determines how easily the crop may be digested by animals; relatively small increases in lignin content may produce large decreases in the digestibility of the crop. Therefore, reducing lignin content would enhance digestibility, facilitating a more efficient use of such crops. In the timber industry, producing wood pulp for papermaking requires the removal of lignin to release the cellulosic content of the timber. The process of removing the lignin consumes large amounts of energy and produces environmentally harmful lignin waste liquors which must be treated prior to disposal. It has also been suggested that residual lignin in paper pulp may produce toxic polychlorinated biphenols when the lignin interacts with chlorine used in the bleaching process. Thus, decreasing lignin content in wood products would be advantageous for papermaking. On the other hand, increasing the lignin content of timber offers the possibility of increased wood strength.
Accordingly, modification of quality and quantity of lignin in plants has been a long-standing interest among breeders and, more recently, among molecular biologists. Recent molecular approaches towards methods for reducing lignin content in plants are typified by: U.S. Pat. No. 5,451,514, "Modification of Lignin Synthesis in Plants"; Canadian Patent No. 2,005,597, "Plants Having Reduced Lignin or Lignin of Altered Quality"; and International Patent Application Publication No. WO 94/23044.
Lignin is a complex polymer of three cinnamyl alcohols, p-coumaryl, coniferyl and sinapyl, all products of phenylpropanoid metabolism. Depending on the plant species or tissue, the relative proportion of the different monomers in lignin can vary significantly. In gymnosperms for example, lignin is predominantly composed of coniferyl alcohol monomer units, whereas angiosperms have significant proportions of sinapyl moieties. The metabolism of lignin production involves many intermediates, enzymatic pathways and, correspondingly, genes. Accordingly, there are several gene/enzyme targets that might be selected to manipulate lignin production through genetic engineering.
Alteration of lignin levels by antisense and sense suppression of gene expression has already been attempted for several enzymes in the phenylpropanoid pathway including PAL (Elkind et al. 1990), CAD (Schuch 1993; Canadian patent 2,005,597; U.S. Pat. No. 5,541,514), 4CL (Lee and Douglas 1994) and COMT (WO 94/23044). However, all of these attempts to modify lignin synthesis are directed at early stages in the synthetic pathway and are therefore likely to interfere with other metabolic processes which share these intermediate steps. It is clear, for example, that interference with early steps in the phenylpropanoid pathway can have undesirable pleiotropic effects (Elkind et al., 1990). In addition, modulating biosynthetic enzymes that act early in the pathway may not be effective because alternative synthetic routes may be available. A better approach to modulating lignin synthesis would be to regulate later stages in the lignin biosynthesis pathway: this would minimize or avoid pleiotropic effects and would likely provide a greater degree of effective control.
The present invention is directed towards the identification of a gene that regulates a later step in the lignin biosynthesis pathway, and the use of this gene to modify lignin content in plants.